Innate immune responses to pathogens are mainly orchestrated by monocytes, macrophages, granulocytes and dendritic cells, which act as a first line of defense against invading microorganisms [1]. Discrimination between self and non self relies on host proteins equipped with the ability to recognize molecular patterns present on foreign organisms [2] [3]. One major family or proteins are the Toll like receptors (TLR), also referred to as pattern recognition receptors (PRR).
TLR4 recognizes lipopolysaccharide (LPS) from Gram negative bacteria, through an extracellular domain characterized by leucine-rich repeats [4], [5], [6]. After its binding to LPS-binding protein (LBP), LPS is transported to a receptor complex involving CD14, TLR4 and an adaptor molecule MD2. Signal transduction is initiated by the interaction of the TIR domain (Toll/IL-1 receptor) of TLR4 with MyD88 (myeloid differentiation primary response protein 88) [7]. A MyD88-independent pathway has been elucidated for TLR4, involving proteins such as Toll-IL1R domain-containing adaptor protein inducing IFN beta (TRIF), TRIF-related adaptor molecules, TANK binding kinase 1 (TANK: TNF receptor associated NFκB kinase) and IκB kinase ε, which then leads to induction of type 1 IFN via interferon regulatory factor 3 (IRF3) [8], [9], [10], [11].
Genetic polymorphisms, for the most part single nucleotide polymorphisms (SNP), are common variants within a population that are found at a frequency of over 1%. They may alter the amino acid sequence (non synonymous SNP), affect the promoter or be silent. Two SNP have been identified within the promoter region of CD14 and one of them, Cys159Thr is related to the incidence and mortality of septic shock [12].
More importantly, SNP in the extracellular domain of TLR4 have been largely investigated. Two co-segregating SNP within the gene encoding TLR4 exhibiting allele frequencies greater than 2% are Asp299Gly and Thr399Ile [13]. These mutations were found in about 10% (0-19%) of control white individuals in most studies. The functional relevance of SNP Asp299Gly has been investigated in hyporesponsiveness to inhaled LPS and allergic asthmatics [13], on the incidence and course of septic shock and various infectious diseases or pathogenesis of atherosclerosis. Accumulating but controversial data point to a role of such SNP TLR4 in altered susceptibility or in the course of these inflammatory or infectious diseases (including allograft reactions, diabetes, coronary stenosis) (reviewed in [14]). One mutated allele is required for the dysfunction [13]. Among 91 people with septic shock during Gram neg. bacteria infection, 4 displayed the Asp299Gly SNP (but not the Thr399Ile) while 0 SNP was detected in the control group [15]. Colonization of pregnant women with Gardnerella spp and other Gram neg commensals was increased by 10 fold in women carrying the mutated Thr399Ile TLR4 allele [16]. A clear correlation was found between severe respiratory syncitial virus (RSV) bronchiolitis in infants and both TLR4 SNP while no correlation with CD14 SNP was found [17].
Blocking of TLR4 may have a beneficial effect on atherosclerosis in mice and two studies found a protective effect of the TLR4 variants on acute coronary events [18], [19], [20]. Rare SNP of TLR4 might also be relevant in some cases such as meningococcemia where Asp299Gly/Thr399Ile were not significant (extracellular region and C terminal domain in the TIR region, [21], [22]).
Functional assays using whole PBMC or monocytes aimed at detecting dysfunctional TLR4 may not be relevant [23], [24], [25].
The association of the SNP with susceptibility to infectious diseases can thus only be regarded as preliminary since studies reporting positive correlations were done with small populations. The finding that the presence of SNP does not influence the activation of monocytes or whole blood by its ligands is further evidence against the hypothesis of a potential role for these SNP in infectious disease susceptibility.
A role for TLR4 polymorphism and cancer has been recently addressed because chronic inflammation may expose to higher risk of tumourigenesis. A TLR4 SNP at position 11 381 was found in Sweden to be more frequent (24.1% versus 19.7%, p=0.02) among people with prostate cancer [26]. A more recent study also concluded that inherited polymorphisms of the innate immune gene TLR4 are associated with risk of prostate cancer [27]. Homozygosity for the variant alleles of eight SNPs was associated with a statistically significantly lower risk of prostate cancer (TLR4—1893, TLR4—2032, TLR4—2437, TLR4—7764, TLR4—11912, TLR4—16649, TLR4—17050, and TLR4—17923), but the TLR4—15844 polymorphism corresponding to 11381G/C was not associated with prostate cancer (GG versus CG/CC: OR, 1.01; 95% confidence interval, 0.79-1.29). Six common haplotypes (cumulative frequency, 81%) were observed; the global test for association between haplotypes and prostate cancer was statistically significant [chi(2)=14.8 on 6 degrees of freedom; P=0.02]. Two common haplotypes were statistically significantly associated with altered risk of prostate cancer.
The first demonstration of a modulatory role for TLR4 in chronic lung inflammation and tumourigenesis has been brought up by Bauer A K et al. [28]. To determine the role of TLR4 in chronic lung inflammation, they compared lung permeability, leukocyte infiltration, and nuclear factor kappa B (NFkappaB) and activator protein 1 (AP-1) DNA binding in butylated hydroxytoluene (BHT)-treated inbred mouse strains with functional TLR4 (OuJ and BALB) and mutated TLR4 [HeJ and BALB(Lps-d)]. They also measured primary tumour formation in these mice after single-carcinogen injection (3-methylcholanthrene) followed by BHT treatment. Mice with functional TLR4 had reduced lung permeability, leukocyte inflammation, and primary tumour formation (BALB(Lps-d), mean=22.3 tumours/mouse, versus BALB, mean=13.9 tumours/mouse, difference=8.4 tumours/mouse, 95% confidence interval=4.6 to 12.1 tumours/mouse; P=0.025) compared with mice with mutated TLR4. NFkappaB DNA binding activity was higher in OuJ than in HeJ mice; however, AP-1 activity was elevated in HeJ mice.
More recently, Okamoto et al. [29], described a correlation between the absence of TLR-4 expression in patients with head and neck cancer and a decreased response to a particular therapeutic treatment comprising the administration of OK-PSA or OK-432, optionally together with UFT (tegafur: uracil, 1:4), and optionally together with a radiotherapy. This document however does not describe or suggest a method according to the present invention of assessing the sensitivity of a subject to a treatment of cancer consisting in a chemotherapeutic treatment of cancer.
Combating cancer efficiently relies on pharmaceutical compounds directly targeting tumour cells or boosting host defense against said cells. Although several anti-cancer therapies are proposed, amongst which feature chemotherapy [anthracyclines, such as doxycycline (DOX), oxali-platinum (herein called PLAT) and cis-platinum (herein called PLAT) are considered as the most efficient cytotoxic agents of the oncologist armamentarium] and radiotherapy [X-Rays (XR)], the benefits of said treatments still tends to be insufficient. Since anthracyclines, oxaliplatinum, cis-platinum and X-Rays represent the basis of up to 70% of anti-cancer therapies, detection of dysfunctions responsible for a reduced response to said treatments appears critical for patient management.